ABSTRACT The goal of this competitive renewal is to identify and validate novel translatable metabolic imaging biomarkers of response to advanced clinically-relevant therapies that target isocitrate dehydrogenase 1 (IDH1) in the treatment of mutant IDH1 glioma. The IDH1 mutation is a `driver mutation' that is present in 70-90% of low- grade glioma and secondary upgraded glioblastoma. In an effort to improve the treatment of mutant IDH1 tumors, novel therapeutic approaches are been developed, and data shows that siRNA targeting both wild-type (wt) and mutant IDH1, as well as new dual (wt/mutant) IDH inhibitors now in clinical trials, lead to a clear response. However, no noninvasive imaging methods are available to assess drug-target engagement or predict response. During the first funding period of this grant we identified several mutant IDH1-driven 1H magnetic resonance spectroscopy (MRS)-detectable metabolic alterations, and developed novel translational hyperpolarized 13C MRS-based imaging approaches that inform on the presence of the mutation. Our preliminary data indicate that response to treatment with emerging dual inhibitors is associated with a reversal of all the 1H MRS-detectable metabolic alterations observed in mutant cells but not with all of the associated hyperpolarized 13C MRS-detectable metabolic fluxes, likely reflecting inhibition of wt IDH1. In addition, levels of glutathione (GSH) and the ratio of GSH to its oxidized form GSSG drop, pointing to additional imageable metabolic reactions associated with response. We therefore hypothesize that using some of our previously identified MRS biomarkers of mutant IDH1, as well as imaging biomarkers of redox status, it will be possible to monitor the therapeutic effects of dual IDH inhibitors that are entering the clinic. Aim 1. To identify 1H and hyperpolarized 13C MRS-detectable metabolic biomarkers associated with response to novel IDH-targeting therapies in mutant IDH1 cells. We will investigate genetically-engineered and patient-derived cell models, and use 1H and hyperpolarized 13C MRS to identify the imageable metabolic alterations that are uniquely associated with response to therapy as determined by inhibition in cell proliferation and/or clonogenic potential. Aim 2. To mechanistically validate 1H and hyperpolarized 13C MRS imaging biomarkers of response. We will use a range of biochemical, cell, and molecular biological assays as well as specific inhibitors to determine the mechanistic link between drug action and the metabolic imaging biomarkers identified in Aim 1. Aim 3. To confirm in vivo the 1H and hyperpolarized 13C MRS metabolic imaging biomarkers as indicators of tumor response to novel IDH-targeting therapies in mutant IDH1 tumors. We will treat tumor-bearing mice, and use MRI with 1H and hyperpolarized 13C MRSI, to longitudinally monitor the effect of therapy on tumor growth and metabolism, and assess the value of our metabolic imaging biomarkers for prediction of response as assessed by tumor size and/or animal survival.